Unraveling the mechanism of action of a new type III secretion system effector AexU from Aeromonas hydrophila

We recently characterized a T3SS effector, AexU, from a diarrheal isolate SSU of Aeromonas hydrophila, which exhibited ADP-ribosyltransferase (ADPRT) activity. Here we provided evidence that AexU also possessed GTPase-activating protein (GAP) activity, which was mainly responsible for host cell apoptosis and disruption of actin filaments. Earlier, we showed that the Δ aexU null mutant was attenuated in a mouse model, and we now demonstrated that while the parental A. hydrophila strain could be detected in the lung, liver, and spleen of infected mice, the Δ aexU mutant was rapidly cleared from these organs resulting in increased survivability of animals. Further, AexU prevented phosphorylation of c-Jun, JNK and IκBα and inhibited IL-6 and IL-8 secretion from HeLa cells. Our data indicated that AexU operated by inhibiting NF-κB and inactivating Rho GTPases. Importantly, however, when the Δ aexU null mutant was complemented with the mutated aexU gene devoid of ADPRT and GAP activities, a higher mortality rate in mice with concomitant increase in the production of pro-inflammatory cytokines/chemokines was noted. These data indicated that either such a mutated AexU is a potent inducer of them or that AexU possesses yet another unknown activity that is modulated by ADPRT and GAP activities and results in this aberrant cytokine/chemokine production responsible for increased animal death.